SUMMARY Regulation of telomerase activity and telomere homeostasis is fundamentally important in growth control. New data in this proposal suggest that human telomerase activity is regulated by ubiquitylation. Specifically, the catalytic subunit of telomerase, hTERT, is post-translationally modified by ubiquitin and directed to the 26S proteasome for degradation. A specific E3 ligase gene has been identified and cloned from humans. This ligase is identical to a well conserved and ancient gene called makorin-1, the founding member of a recently discovered gene family of unknown function. MKRN1 encodes a RING-zinc finger protein that catalyzes the ligation of ubiquitin to hTERT. High levels of MKRN1 expression in cancer cells lead to telomere DNA erosion and a reduction in telomerase activity as measured by the TRAP assay. In order to understand this new pathway for hTERT regulation, a series of objectives are planned to examine in detail the consequences of over and under-expressing MKRN1 in cancer cells. The ultimate goal of this research is to identify novel targets for therapeutic intervention in cancer and to explore how the proteasome pathway intersects mechanistically with the telomere homeostatic pathway. Telomeres are the physical ends of chromosomes encrypted by a repeat DNA sequence (TTAGGG) and maintained by telomerase. We have new evidence that human telomerase is regulated by the ubiquitin/proteasome pathway and we have cloned a telomerase killer gene called MKRN1. Since most tumor cells inappropriately express telomerase, a telomerase inactivation pathway is of clinical significance in designing new ways to intervene in cancer. [unreadable] [unreadable] [unreadable]